Development and evaluation of a meat mitochondrial metagenomic (3MG) method for composition determination of meat from fifteen mammalian and avian species

Abstract Background Bioassessment and biomonitoring of meat products are aimed at identifying and quantifying adulterants and contaminants, such as meat from unexpected sources and microbes. Several methods for determining the biological composition of mixed samples have been used, including metabarcoding, metagenomics and mitochondrial metagenomics. In this study, we aimed to develop a method based on next-generation DNA sequencing to estimate samples that might contain meat from 15 mammalian and avian species that are commonly related to meat bioassessment and biomonitoring. Results In this project, we found the meat composition from 15 species could not be identified with the metabarcoding approach because of the lack of universal primers or insufficient discrimination power. Consequently, we developed and evaluated a meat mitochondrial metagenomics (3MG) method. The 3MG method has four steps: (1) extraction of sequencing reads from mitochondrial genomes (mitogenomes); (2) assembly of mitogenomes; (3) mapping of mitochondrial reads to the assembled mitogenomes; and (4) biomass estimation based on the number of uniquely mapped reads. The method was implemented in a python script called 3MG. The analysis of simulated datasets showed that the method can determine contaminant composition at a proportion of 2% and the relative error was < 5%. To evaluate the performance of 3MG, we constructed and analysed mixed samples derived from 15 animal species in equal mass. Then, we constructed and analysed mixed samples derived from two animal species (pork and chicken) in different ratios. DNAs were extracted and used in constructing 21 libraries for next-generation sequencing. The analysis of the 15 species mix with the method showed the successful identification of 12 of the 15 (80%) animal species tested. The analysis of the mixed samples of the two species revealed correlation coefficients of 0.98 for pork and 0.98 for chicken between the number of uniquely mapped reads and the mass proportion. Conclusion To the best of our knowledge, this study is the first to demonstrate the potential of the non-targeted 3MG method as a tool for accurately estimating biomass in meat mix samples. The method has potential broad applications in meat product safety.

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Flesh ID: Nanopore Sequencing Combined with Offline BLAST Search for the Identification of Meat Source

Foods ◽

10.3390/foods9101392 ◽

2020 ◽

Vol 9 (10) ◽

pp. 1392

Author(s):

Jonas Kissenkötter ◽

Susanne Böhlken-Fascher ◽

Ahmed Abd El Wahed

Keyword(s):

Gold Standard ◽

Animal Species ◽

Detection System ◽

Meat Product ◽

Meat Products ◽

Nanopore Sequencing ◽

Pork Meat ◽

Blast Search ◽

International Regulations ◽

Pcr Assays

Detection of animal species in meat product is crucial to prevent adulterated and unnecessary contamination during processing, in addition to avoid allergy and religious consequences. Gold standard is the real-time PCR assays, which has a limited target capability. In this study, we have established a rapid sequencing protocol to identify animal species within hours. Sequencing was achieved by nanopore sequencing and data analysis via offline BLAST search. The whole procedure was conducted in a mobile suitcase lab. As per national and international regulations, the developed assay detected adulteration of pork meat with 0.1% of horse, chicken, turkey, cattle, sheep, duck, rabbit, goat, and donkey. The developed test could be used on-site as a rapid and mobile detection system to determine contamination of meat products.

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A Simple and Reliable Single Tube Septuple PCR Assay for Simultaneous Identification of Seven Meat Species

Foods ◽

10.3390/foods10051083 ◽

2021 ◽

Vol 10 (5) ◽

pp. 1083

Author(s):

Zhendong Cai ◽

Song Zhou ◽

Qianqian Liu ◽

Hui Ma ◽

Xinyi Yuan ◽

...

Keyword(s):

Dna Sequences ◽

Animal Species ◽

Meat Product ◽

Meat Products ◽

Pcr Assay ◽

Specific Primers ◽

Meat Species ◽

Pcr Method ◽

Species Specific ◽

Simultaneous Identification

Multiplex PCR methods have been frequently used for authentication of meat product adulteration. Through screening of new species-specific primers designed based on the mitochondrial DNA sequences, a septuple PCR method is ultimately developed and optimized to simultaneously detect seven species including turkey (110 bp), goose (194 bp), pig (254 bp), sheep (329 bp), beef (473 bp), chicken (612 bp) and duck (718 bp) in one reaction. The proposed method has been validated to be specific, sensitive, robust and inexpensive. Taken together, the developed septuple PCR assay is reliable and efficient, not only to authenticate animal species in commercial meat products, but also easily feasible in a general laboratory without special infrastructures.

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Combining a COI Mini-Barcode with Next-Generation Sequencing for Animal Origin Ingredients Identification in Processed Meat Product

Journal of Food Quality ◽

10.1155/2020/2907670 ◽

2020 ◽

Vol 2020 ◽

pp. 1-9 ◽

Cited By ~ 2

Author(s):

Yanyi Pan ◽

Deyi Qiu ◽

Jian Chen ◽

Qiaoyun Yue

Keyword(s):

Next Generation Sequencing ◽

Animal Species ◽

Detection Efficiency ◽

Meat Product ◽

Animal Origin ◽

Processed Meat ◽

Next Generation ◽

Universal Primer ◽

Clone Sequencing ◽

Generation Sequencing

For revealing animal species in complex or adulterated processed meat product, we presented a method combining a novel cytochrome oxidase I (COI) mini-barcode with next-generation sequencing (NGS), which identifies various animal species (swine, bovine, Caprinae, and some of fish, shrimp, and poultry) accurately and efficiently in processed meat products. We designed a universal primer based on 140 sequences from 51 edible animal species. A mixture of 12 species raw meat samples were identified with the clone sequencing and also with a mini-barcode- (136 bp) combined NGS method, respectively. The mini-barcode of these 12 species was 100% identical to the target species sequence by Sanger sequencing. Compared to the clone sequencing method, the NGS method is superior in accuracy, sensitivity, and detection efficiency. Various edible animal species were identified in the species level both in the mixed samples and the 7 heavily processed food products. Moreover, some unlabeled species and dubious contamination were detected as well.

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Intraspecific Diversity of Lactobacillus curvatus, Lactobacillus plantarum, Lactobacillus sakei, and Leuconostoc mesenteroides Associated with Vacuum-Packed Meat Product Spoilage Analyzed by Randomly Amplified Polymorphic DNA PCR

Journal of Food Protection ◽

10.4315/0362-028x-69.10.2403 ◽

2006 ◽

Vol 69 (10) ◽

pp. 2403-2410 ◽

Cited By ~ 11

Author(s):

ROSA AZNAR ◽

EMPAR CHENOLL

Keyword(s):

Lactobacillus Plantarum ◽

Leuconostoc Mesenteroides ◽

Meat Product ◽

Meat Products ◽

Storage Period ◽

Intraspecific Diversity ◽

Universal Primers ◽

Lactobacillus Sakei ◽

Randomly Amplified Polymorphic Dna ◽

Lactobacillus Curvatus

The intraspecific diversity of Leuconostoc mesenteroides, Lactobacillus curvatus, Lactobacillus sakei, and Lactobacillus plantarum was analyzed by randomly amplified polymorphic DNA (RAPD) PCR with universal primers M13 and T3. The study included 100 reference strains and 210 isolates recovered from two vacuum-packed Spanish meat products, fiambre de magro adobado and morcilla, previously identified by rDNA–restriction fragment length polymorphism profiles. The RAPDM13 profiles identified isolates at species level in L. plantarum and L. mesenteroides, while RAPD-T3 provided profiles in L. sakei. The combination of RAPD-M13 and RAPD-T3 fingerprints revealed a total of 17 profiles in L. mesenteroides, 6in L. sakei, 12 in L. plantarum, and6in L. curvatus. Of these, six profiles corresponding to L. mesenteroides and one corresponding to L. sakei were found in both products. The Shannon-Weaver diversity index (H′), calculated according to RAPD-M13 and RAPD-T3 profiles during storage, revealed that most profiles appeared only in single samplings in both products, indicating a high strain substitution rate during chilled storage of vacuum-packed meat products. When bloating appeared, only one profile corresponding to L. mesenteroides subsp. dextranicum was present throughout the storage period.

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Proximate Chemical Analysis, Fatty Acid Profile and Microstructural Characteristics of Dromedary Camel Fats (Hump, Renal and Mesentery)

International Journal of Veterinary Science ◽

10.37422/ijvs/20.014 ◽

2020 ◽

Keyword(s):

Fatty Acid ◽

Fatty Acid Profile ◽

Meat Product ◽

Oxidation Stability ◽

Meat Products ◽

Raw Material ◽

Dromedary Camel ◽

Mean Values ◽

Animal Fat ◽

Mesenteric Fat

The quality, safety, and suitability of animal fat for processing of a specific meat product is a critical issue. Increasing the human awareness about the health aspects associated with increased intake of animal fat, makes camel fat a suitable raw material for meat processing due to its excellent nutritional contribution. Therefore, the target of this study is examination of the sensory, physicochemical, fat oxidation, fatty acid profile, and other quality parameters of camel fat to evaluate the feasibility for processing of different meat products. To achieve this goal, 30 fat samples each from the hump, renal, and mesentery of Arabian male camels were investigated. The results showed that both the renal and mesenteric fat had honey color and medium-soft texture, while the hump had greyish-white color and hard texture. The sensory panel scores were significantly different between the hump and other fats. Hump fat had significantly (P<0.05) higher moisture, protein, and collagen content, while higher fat content was recorded in mesenteric fat. The fatty acid analysis showed that hump had high SFA and very low PUFA in comparison with both renal and mesenteric fat. Camel fat had high oxidation stability, and the mean values were very low in comparison with the levels of quality and acceptability. The ultrastructural analysis showed that hump fat had high elastin fibers which increase its hardness. The results indicated that both renal and mesenteric fat were more suitable for the production of various meat products than the hump.

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Development of a Methodology for Estimating the Ergosterol in Meat Product-Borne Toxigenic Moulds to Evaluate Antifungal Agents

Foods ◽

10.3390/foods10020438 ◽

2021 ◽

Vol 10 (2) ◽

pp. 438

Author(s):

Micaela Álvarez ◽

Alicia Rodríguez ◽

Elena Bermúdez ◽

Elia Roncero ◽

María J. Andrade

Keyword(s):

Antifungal Agents ◽

Limit Of Detection ◽

Meat Product ◽

Meat Products ◽

Ergosterol Biosynthesis ◽

Array Detector ◽

Meat Industry ◽

Ergosterol Content ◽

Validation Parameters ◽

Toxigenic Moulds

Antifungal agents are commonly used in the meat industry to prevent the growth of unwanted moulds, such as toxigenic ones, on dry-cured meat products. For enhancing the application of antifungals, their mode of action must be evaluated. Their effect on the mould ergosterol content is one of the most studied ones, since it is the target site of some commercialised antifungals or of those that are in development. The aim of this study was to develop a methodology for determining how the antifungal agents used in the meat industry work. A method for analysing ergosterol was firstly developed using high-performance liquid chromatography with fluorescence detection coupled to a diode array detector (HPLC-FLD/DAD). The chromatographically optimised conditions (gradient and mobile phases) allowed us to reduce the time per analysis with respect to previously published methods up to 22 min. Withing the six checked extraction methods, method 5, showing the best mean recovery values (99.51%), the shortest retention time (15.8 min), and the lowest standard deviation values (9.92) and working temperature (60 °C), was selected. The limit of detection and limit of quantification were 0.03 and 0.1 µg/mL, respectively. All the validation parameters corroborated the method’s suitability. Finally, its feasibility for evaluating the effect of a commercial antifungal preparation (AP) and different herbs that are frequently added to meat products on the ergosterol content of several toxigenic moulds was studied. Differences at the strain level were obtained in the presence of AP. Moreover, the addition of herbs significantly reduced the ergosterol content in Penicillium nordicum up to 83.91%. The developed methodology is thus suitable for screening the antifungals’ role in altering mould ergosterol biosynthesis before their application in real meat products.

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The Effect of the Addition of Fiber Preparations on the Color of Medium-Grounded Pasteurized and Sterilized Model Canned Meat Products

Molecules ◽

10.3390/molecules26082247 ◽

2021 ◽

Vol 26 (8) ◽

pp. 2247

Author(s):

Mirosław Słowiński ◽

Joanna Miazek ◽

Krzysztof Dasiewicz ◽

Marta Chmiel

Keyword(s):

Meat Product ◽

Meat Products ◽

Quality Characteristics ◽

Consumer Acceptance ◽

Meat Industry ◽

Industrial Practice ◽

Yellow Color ◽

Hue Angle ◽

Canned Meat ◽

Color Components

A beneficial aspect of the use of fiber preparations in the meat industry is the improvement of some quality characteristics of meat products. However, the preparation added in the amount of 3 or 6% may affect their color. The effect of the addition of barley, wheat and oat fiber preparations with different fiber lengths, in quantities allowing the product to be indicated as “high in fiber” or “source of fiber”, to pasteurized or sterilized medium-grounded canned meat products on their color, was determined. In the obtained canned meat products, the basic chemical composition and the L*, a* and b*, C* (Chroma) and h* (hue angle) color components were determined. The addition of the barley fiber preparation BG 300 to the model canned meat products caused a significant (p ≤ 0.05) darkening and an increase in the proportion of yellow color. In an industrial practice, this may result in poorer consumer acceptance of the meat product. Fiber length of wheat and barley fiber had no effect on the color components of products. The 6% addition of the wheat fiber preparations WF 200R and WF 600R or the oat fiber preparations HF 200 and HF 600 caused an apparent lightening of their color (ΔE > 2) compared to the control products.

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PSV-25 Detection of heart and aorta tissue peptide markers by the multiple-reaction monitoring method

Journal of Animal Science ◽

10.1093/jas/skaa278.636 ◽

2020 ◽

Vol 98 (Supplement_4) ◽

pp. 362-363

Author(s):

Daniil Khvostov ◽

Natalya Vostrikova ◽

Irina M Chernukha

Keyword(s):

Amino Acid ◽

High Performance ◽

Multiple Reaction Monitoring ◽

Meat Product ◽

Meat Products ◽

Amino Acid Sequences ◽

Composition Analysis ◽

Reaction Monitoring ◽

Russian Science ◽

Monitoring Method

Abstract Functional, particularly personalized meat-based foods are of more in demand by a consumer today. Functional additives, such as plant components and animal proteins from bovine or porcine tissues have been successfully used. With many ingredients added to foods, it is important to provide quality and composition monitoring to confirm the products’ authenticity, to identify undeclared or rarely used types of raw meat in product formulations. For example, if animal heart tissue is a component of a product formulation or if aorta tissue presents in a product due to improper trimming. Different methods are used to identify raw materials, including new approaches in proteomics and peptidomics that are considered the most effective modern methods nowadays. The purpose of the study is meat product composition analysis and special biomarker peptide identification to confirm the presence of heart and aorta tissue in a finished meat product. Over 20 amino acid sequences were checked based on earlier obtained data. Those amino acid sequences were analyzed with a high-performance liquid chromatography with mass spectrometric detection as described. The MS settings were selected using the Skyline. Signal-to-Noise ratio (S/N) over 10 units were used to choose the best peptide candidates. Seven peptides were found in porcine hearts. The best candidate was peptide VNVDEVGGEALGR (S/N - 73.10±5.3) from β-Hemoglobin. Two marker peptides from serum albumin were selected for pork aorta: TVLGNFAAFVQK (S/N 53.51±2.4) and EVTEFAK (S/N 31.69±4.1). These biomarkers showed the best detection and specificity. The multiply reaction monitoring method made it possible to identify the most/best specific peptides—biomarkers that could confirm the heart and/or aorta in meat products. The method can be used for comparative research or identification of best peptides that are specific to any type of animal tissue. The work was supported by the Russian Science Foundation, project no. 16-16- 10073.

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A Multiplex PCR Assay Mediated by Universal Primers for the Detection of Adulterated Meat in Mutton

Journal of Food Protection ◽

10.4315/0362-028x.jfp-18-302 ◽

2019 ◽

Vol 82 (2) ◽

pp. 325-330 ◽

Cited By ~ 1

Author(s):

WANWAN LIU ◽

XIAONAN WANG ◽

JING TAO ◽

BANGSHENG XI ◽

MAN XUE ◽

...

Keyword(s):

Multiplex Pcr ◽

Detection System ◽

Meat Products ◽

Pcr Primers ◽

Rapid Identification ◽

Food Samples ◽

Detection Sensitivity ◽

Universal Primers ◽

Multiplex Pcr Assay ◽

Multiple Pcr

ABSTRACT This study aimed to establish a multiplex PCR detection system mediated by “universal primers,” which would be able to determine whether mutton meat contained nonmutton ingredients from rats, foxes, and ducks. Based on the sequence variation of specific mitochondrial genes, nine different multiplex PCR primers were designed, and four kinds of meat products were rapidly identified by electrophoresis using an optimized multiplex PCR system based on the molecular weight differences of the amplified products. Multiplex PCR applications optimized for meat food source from food samples for testing was used to verify the accuracy of the identification method. The results showed that the primers in multiple PCR system mediated by universal primers could be used for the rapid identification of rat, fox, duck, and sheep meat in mutton products, and the detection sensitivity could reach 0.05 ng/μL. The identification of food samples validated the practical value of this method. Therefore, a multiplex PCR system mediated by universal primers was established, which can be used to quickly identify the origin of animal ingredients from rats, foxes, and ducks in mutton products.

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Research on Modified Atmosphere Packaging in Fresh Red Meat Product Marketing

Applied Mechanics and Materials ◽

10.4028/www.scientific.net/amm.200.351 ◽

2012 ◽

Vol 200 ◽

pp. 351-355

Author(s):

Peng Zhao ◽

Lei Cao

Keyword(s):

Storage Temperature ◽

Operation Mode ◽

Meat Product ◽

Modified Atmosphere Packaging ◽

Meat Products ◽

Red Meat ◽

Modified Atmosphere ◽

Mixed Gas ◽

Gas Barrier ◽

Barrier Materials

According to the national conditions in China, this article is aiming at introducing the application statues of the modified atmosphere packaging of the fresh red meat product. It has expounded in detail the product feature of the fresh red meat and the mechanization of retaining freshness in the way of the modified atmosphere packaging. From four aspects, this article analyzes the influencing factors of the fresh red meat, such as the proportion of mixed gas, gas barrier materials, meat splitting ways and the meat storage temperature. In accordance with the market sales environment, it has discussed the specific improvement operation mode in quality of meat, the handle ability and the effect of packing, as well as the problems which should be noticed in this process. At the end, it has look into the promising future in the modified atmosphere packaging of fresh red meat products.

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